PGEX-2T, 25 µg

Simplified package with only the pGEX vector in the pack (E. coli BL21 will continue to be available separately, code no 27-1542-01. Updated Product Specification sheets. Maintained product quality and quantity. New code numbers.

Thirteen pGEX vectors are available (see Figure). Nine of the vectors have an expanded multiple cloning site (MCS) that contains six restriction sites. The expanded MCS facilitates the unidirectional cloning of cDNA inserts obtained from libraries constructed using many available lambda vectors. pGEX-6P-1, pGEX-6P-2, and pGEX-6P-3 each encode the recognition sequence for site-specific cleavage by PreScission Protease, (see PreScission Protease) between the GST domain and the multiple cloning site. pGEX-4T-1, pGEX-4T-2, and pGEX-4T-3 are derived from pGEX-2T and contain a Thrombin recognition site. pGEX-5X-1, pGEX-5X-2, and pGEX5X-3 are derivatives of pGEX-3X and possess a Factor Xa recognition site.

pGEX-2TK is designed to allow the detection of expressed proteins by directly labeling the fusion products in vitro (1). This vector contains the recognition sequence for the catalytic subunit of cAMP-dependent protein kinase obtained from heart muscle. The protein kinase site is located between the GST domain and the MCS. Expressed proteins can be directly labeled using protein kinase and [gamma-32P]ATP and readily detected using standard radiometric or autoradiographic techniques. pGEX-2TK is a derivative of pGEX-2T; its fusion proteins can be cleaved with Thrombin.

Cleavage of pGEX-6P GST fusion proteins occurs between the Gln and Gly residues of the recognition sequence Leu-Glu-Val-Leu-Phe-Gln-Gly-Pro (2). Low temperature (5°C) digestion minimizes the degradation of the protein of interest. Because PreScission Protease has been engineered with a GST tag, it can also be removed from the cleavage mixture simultaneously with the GST portion of the fusion protein. The pGEX-6P Expression Vectors permit convenient site-specific cleavage and simultaneous purification on Glutathione Sepharose. The pGEX-6P series provides all three translational reading frames linked between the GST coding region and the multiple cloning site.

PGEX-2T, 25 µg

The Glutathione S-transferase (GST) Gene Fusion System from GE Healthcare is a
versatile system for the expression, purification, and detection of GST-tagged
proteins produced in E. coli. The system consists of three major components:
pGEX plasmid expression vectors, products for GST purification, and a variety
of GST detection products. A series of site-specific proteases for cleavage of
the GST tag complements the system. The GST affinity tag permits a mild
purification process that does not affect a protein’s native structure and
function.
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• Simplified package with only the pGEX vector in the pack (E. coli BL21 will
continue to be available separately, code no 27-1542-01)
• Updated Product Specification sheets
• Maintained product quality and quantity
• New code numbers
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This handbook describes the Glutathione S-transferase (GST) Gene Fusion System,
a versatile system for the expression, purification and detection of fusion
proteins produced in Escherichia coli. Covering topics from cloning procedures
to removal of the GST tag by enzymatic cleavage and including a helpful
troubleshooting section, this handbook is a practical guide.
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Almost all protein samples need further preparation after collection. The
quality of sample preparation is crucial to get the best possible analytical
results. This selection guide helps you to find the appropriate methods and
products to obtain proteins for further studies in scales from nanograms to
milligrams of purified protein. Three separate workflows are available for
different sources of target protein: Recombinant proteins, antibodies, and
proteins from natural sources. These workflows are further refined and adapted
to suit different number of samples, from a few to many, such as in screening
applications.
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Map of the glutathione S-transferase fusion vectors showing the reading frames
and main features. Even though stop codons in all three frames are not depicted
in this map, all thirteen vectors have stop codons in all three frames
downstream from the multiple cloning site.
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